This proposal seeks to develop novel instrumentation for in vivo fluorescence microscopy based on micro-optical probes that are capable of imaging biological cells deep within tissues of live animals or humans in a minimally invasive manner. Capitalizing on recent advances in telecom micro-optics, compound micro-optical probes as small as 350-1000 mu/m in diameter will provide micron-scale resolution of detail. This technology for in vivo micro-imaging will impact several broad classes of applications, including development of novel drugs and therapeutic methods, minimally invasive surgical procedures, and biomedical research on live animal models of disease. Specific aims are to: (1) Develop a second generation of micro-optical instrumentation providing multi-functional capabilities for in vivo imaging in deep tissues previously inaccessible to bulky microscope optics. In particular, we will: Design new micro-optical probes that are smaller, enable higher resolution imaging, and provide broader fields of view than those currently available for micro-optics based microscopy; Equip micro-optical probes with capabilities for fluidic delivery and sampling, enabling in vivo drug delivery to the imaging field and off-line genetic, biochemical, or cytometric analysis of fluid specimens; Equip micro-optical probes with electrodes for making in vivo electrophysiological measurements at the imaging field. Further aims are to: (2) Develop high-speed micro imaging for tracking fast cellular dynamics. Demonstration of this capability in vivo will involve imaging of neuronal Ca 2+ dynamics; (3) Construct a high-speed image stabilization mechanism for keeping micro-optical or conventional microscopy images in focus while tissues are moving in anesthetized or alert subjects. Such capabilities will improve the quality of micro-imaging data and greatly expand the scope of applicability. To validate each new micro-imaging methodology in vivo, we will study neurons, dendrites, and dendritic spines in the hippocampal brain area of live rodents, where in vivo cellular imaging has previously been prohibitive.